Predicting responses to inhibitory synaptic input in substantia nigra pars reticulata neurons.

The changes in firing probability produced by a synaptic input are usually visualized using the poststimulus time histogram (PSTH). It would be useful if postsynaptic firing patterns could be predicted from patterns of afferent synaptic activation, but attempts to predict the PSTH from synaptic potential waveforms using reasoning based on voltage trajectory and spike threshold have not been successful, especially for inhibitory inputs. We measured PSTHs for substantia nigra pars reticulata (SNr) neurons inhibited by optogenetic stimulation of striato-nigral inputs or by matching artificial inhibitory conductances applied by dynamic clamp. The PSTH was predicted by a model based on each SNr cell's phase-resetting curve (PRC). Optogenetic activation of striato-nigral input or artificial synaptic inhibition produced a PSTH consisting of an initial depression of firing followed by oscillatory increases and decreases repeating at the SNr cell's baseline firing rate. The phase resetting model produced PSTHs closely resembling the cell data, including the primary pause in firing and the oscillation. Key features of the PSTH, including the onset rate and duration of the initial inhibitory phase, and the subsequent increase in firing probability could be explained from the characteristic shape of the SNr cell's PRC. The rate of damping of the late oscillation was explained by the influence of asynchronous phase perturbations producing firing rate jitter and wander. Our results demonstrate the utility of phase-resetting models as a general method for predicting firing in spontaneously active neurons and their value in interpretation of the striato-nigral PSTH. NEW & NOTEWORTHY The coupling of patterned presynaptic input to sequences of postsynaptic firing is a Gordian knot, complicated by the multidimensionality of neuronal state and the diversity of potential initial states. Even so, it is fundamental for even the simplest understanding of network dynamics. We show that a simple phase-resetting model constructed from experimental measurements can explain and predict the sequence of spike rate changes following synaptic inhibition of an oscillating basal ganglia output neuron.

Contributions of Kv7-mediated potassium current to sub- and suprathreshold responses of rat layer II/III neocortical pyramidal neurons.

After block of Kv1- and Kv2-mediated K(+) currents in acutely dissociated neocortical pyramidal neurons from layers II/III of rat somatosensory and motor cortex, the remaining current is slowly activating and persistent. We used whole cell voltage clamp to show that the Kv7 blockers linopirdine and XE-991 blocked a current with similar kinetics to the current remaining after combined block of Kv1 and Kv2 channels. This current was sensitive to low doses of linopirdine and activated more slowly and at more negative potentials than Kv1- or Kv2-mediated current. The Kv7-mediated current decreased in amplitude with time in whole cell recordings, but in most cells the current was stable for several minutes. Current in response to a traditional M-current protocol was blocked by muscarine, linopirdine, and XE-991. Whole cell slice recordings revealed that the Q10 for channel deactivation was ∼2.5. Sharp electrode current-clamp recordings from adult pyramidal cells demonstrated that block of Kv7-mediated current with XE-991 reduced rheobase, shortened the latency to firing to near rheobase current, induced more regular firing at low current intensity, and increased the rate of firing to a given current injection. XE-991 did not affect single action potentials or spike frequency adaptation. Application of XE-991 also eliminated subthreshold voltage oscillations and increased gain for low-frequency inputs (<10 Hz) without affecting gain for higher frequency inputs. These data suggest important roles for Kv7 channels in subthreshold regulation of excitability, generation of theta-frequency subthreshold oscillations, regulation of interspike intervals, and biasing selectivity toward higher frequency inputs.

Functional roles of Kv1 channels in neocortical pyramidal neurons.

Pyramidal neurons from layers II/III of somatosensory and motor cortex express multiple Kv1 alpha-subunits and a current sensitive to block by alpha-dendrotoxin (alpha-DTX). We examined functional roles of native Kv1 channels in these cells using current-clamp recordings in brain slices and current- and voltage-clamp recordings in dissociated cells. alpha-DTX caused a significant negative shift in voltage threshold for action potentials (APs) and reduced rheobase. Correspondingly, a ramp-voltage protocol revealed that the alpha-DTX-sensitive current activated at subthreshold voltages. AP width at threshold increased with successive APs during repetitive firing. The steady-state threshold width for a given firing rate was similar in control and alpha-DTX, despite an initially broader AP in alpha-DTX. AP voltage threshold increased similarly during a train of spikes under control conditions and in the presence of alpha-DTX. alpha-DTX had no effect on input resistance or resting membrane potential and modest effects on the amplitude or width of a single AP. Accordingly, experiments using AP waveforms (APWs) as voltage protocols revealed that alpha-DTX-sensitive current peaked late during the AP repolarization phase. Application of alpha-DTX increased the rate of firing to intracellular current injection and increased gain (multiplicative effects), but did not alter spike-frequency adaptation. Consistent with these findings, voltage-clamp experiments revealed that the proportion of outward current sensitive to alpha-DTX was highest during the interval between two APWs, reflecting slow deactivation kinetics at -50 mV. Finally, alpha-DTX did not alter the selectivity of pyramidal neurons for DC versus time-varying stimuli.

Presynaptic effects of group III metabotropic glutamate receptors on excitatory synaptic transmission in the retina.

Metabotropic glutamate receptors (mGluRs) are located in both plexiform layers in the retina and may modulate transmission between photoreceptors and bipolar cells and between bipolar and ganglion cells. We investigated whether mGluR activation modulates excitatory synaptic input to bipolar cells and ganglion cells in the salamander retinal slice preparation. The group III mGluR agonist L-2-amino-4-phosphonobutyric acid (AP4) inhibited monosynaptic excitatory postsynaptic currents (EPSCs) in ganglion cells evoked by electrical stimuli, whereas group I and group II agonists had no significant effect. AP4 reduced the frequency but not the amplitude of ganglion cell miniature EPSCs, suggesting a presynaptic action at bipolar cell terminals. AP4 also reduced ganglion cell EPSCs evoked by the offset of a light stimulus, suggesting that group III mGluRs modulate release from OFF bipolar cells. Comparison of light-evoked EPSCs in OFF bipolar cells and ganglion cells indicated that AP4 reduced ganglion cell EPSCs by acting primarily at bipolar cell terminals, and to a lesser extent at photoreceptor terminals. The group II/III mGluR antagonist (RS)-alpha-cyclopropyl-4-phosphonophenylglycine (CPPG) blocked the effect of AP4 at bipolar cell terminals, consistent with localization of group III mGluRs at these sites. However, CPPG did not increase EPSCs at light offset, indicating that activation of group III mGluRs by synaptic glutamate does not play a large role in modulating transmission from bipolar cells to ganglion cells.

Morphological and electrophysiological evidence for an ionotropic GABA receptor of novel pharmacology.

Evidence from toxicological studies suggested that an ionotropic GABA receptor of novel pharmacology (picrotoxin-insensitive, bicuculline-sensitive) exists in the chick embryo retina. In this report, we provide direct morphological and electrophysiological evidence for the existence of such an iGABA receptor. Chick embryo retinas (14-16 days old) incubated in the presence of kainic acid showed pronounced histopathology in all retinal layers. Maximal protection from this toxicity required a combination of bicuculline and picrotoxin. Individual application of the antagonists indicated that a picrotoxin-insensitive, bicuculline-sensitive GABA receptor is likely to be present on ganglion and amacrine, but not bipolar, cells. GABA currents in embryonic and mature chicken retinal neurons were measured by whole cell patch clamp. GABA was puffed at the dendritic processes in the IPL. Picrotoxin (500 microM, in the bath) eliminated all (>95%) the GABA current in the majority of ganglion and amacrine cells tested, but many cells possessed a substantial picrotoxin-insensitive component. This current was eliminated by bicuculline (200 microM). This current was not a transporter-associated current, since it was not altered by GABA transport blockers or sodium removal. The current-voltage relation was linear and reversed near E(Cl), as expected for a ligand-gated chloride current. Both pentobarbital and lorazepam enhanced the picrotoxin-insensitive current. We conclude that chicken retinal ganglion and amacrine cells express a GABA receptor that is GABA-A-like, in that it can be blocked by bicuculline, and positively modulated by barbiturates and benzodiazepines, but is insensitive to the noncompetitive blocker picrotoxin. Understanding the molecular properties of this receptor will be important for understanding both physiological GABA neurotransmission and the pathology of GABA receptor overactivation.

AMPA receptor kinetics limit retinal amacrine cell excitatory synaptic responses.

Amacrine cells that respond transiently to maintained illumination are thought to mediate transient inhibitory input to ganglion cells. The excitation of these transient amacrine cells is thought to be limited by inhibitory feedback to bipolar cells. We investigated the possibility that desensitizing AMPA and/or kainate (KA) receptors on amacrine cells might also limit the duration of amacrine cell excitation. To determine how these receptors might affect amacrine cell input and output, we made whole-cell recordings from amacrine and ganglion cells in the salamander retinal slice. The specific AMPA receptor antagonist GYKI-53655 blocked non-NMDA receptor-mediated amacrine cell excitatory postsynaptic currents (EPSCs) and kainate puff-elicited currents, indicating that AMPA, and not KA, receptors mediated the responses. Cyclothiazide, an agent that reduces AMPA receptor desensitization, increased the amplitude and duration of amacrine cell EPSCs. To measure the output of transient amacrine cells, we recorded glycinergic inhibitory postsynaptic currents (IPSCs) from ganglion cells, and found that these were also enhanced by cyclothiazide. Thus, prolongation of amacrine cell AMPA receptor activation enhanced amacrine cell output. Current responses elicited by puffing glycine onto ganglion cell dendrites were not affected by cyclothiazide, indicating that the enhancement of glycinergic IPSCs was not due to a direct effect on glycine receptors. These data suggest that rapid AMPA receptor desensitization and/or deactivation limits glycinergic amacrine cell excitation and the resulting inhibitory synaptic output.

Glutamate uptake limits synaptic excitation of retinal ganglion cells.

EPSCs of retinal ganglion cells decay more slowly than do those of most other CNS neurons, in part because of the long time course of glutamate release from bipolar cells. Here we investigated how glutamate clearance and AMPA receptor desensitization affect ganglion cell EPSCs in the salamander retinal slice preparation. Inhibition of glutamate uptake greatly prolonged ganglion cell EPSCs evoked by light or monosynaptic electrical stimuli but had little effect on spontaneous miniature EPSCs (mEPSCs). This suggests that single quanta of glutamate are cleared rapidly by diffusion but multiple quanta can interact to lengthen the postsynaptic response. Some interaction between quanta is likely to occur even when glutamate uptake is not inhibited. This seems to depend on quantal content, because reducing glutamate release with low Ca2+, paired-pulse depression, or weak stimuli shortened the EPSC decay. High quantal content glutamate release may lead to desensitization of postsynaptic receptors. We reduced the extent of AMPA receptor desensitization by holding ganglion cells at positive potentials. This increased the amplitude of the late phase of evoked EPSCs but did not affect the decay rate after the first 50 msec of the response. In contrast, the holding potential had little effect on mEPSC kinetics. Our results suggest that desensitization limits the late phase of AMPA receptor-mediated EPSCs, whereas glutamate uptake controls the duration of both AMPA and NMDA receptor-mediated responses.